Pancreatic cancer is a type of tumor with high mortality rate (˜99.9%, for definitive diagnosed patients). It was estimated that in USA, in 2005, there were 32,180 incidence cases of pancreatic cancer, accounting for 2% of all incidence cancer cases; 31,800 death cases, ranking the 4th and 5th in the male and female respectively among all causes of deaths related to cancer, taking 5%-6% of all causes of deaths related to cancer. In EU, it was estimated that in 2002, there were 55,100 incidence cases and 59,300 death cases of pancreatic cancer, without definite variations in different genders and races and commonly the prognosis is poor. A statistics for the global oncoming and death of pancreatic cancer patients is shown in Table 1, wherein the term “Incidence Cases” refers to the number of pancreatic cancer cases newly diagnosed in 2002, and the term “Death Cases” refers to the population that were diagnosed as pancreatic cancer by 2002 and died in 2002.
TABLE 1A statistics of global pancreatic cancer oncoming and death in 2002MaleFemaleIncidenceIncidenceDeathCasesDeath CasesCasesCasesUSA15,90015,00015,80015,400EU28,40029,60026,70029,700Others80,60075,00065,00062,400
Therefore, seeking markers for pancreatic cancer and detecting the pancreatic cancer with high accuracy have become an extremely and important precondition of early stage diagnosis and treatment for pancreatic cancer. Although more and more disease markers have been discovered and applied in the clinically general survey and diagnosis of diseases and in therapeutic efficacy monitoring, there are still significant deficiencies in their clinical application. For instance, the tumor markers alpha-fetoprotein (AFP), lactic dehydrogenase (LHD) and carcinoembryonic antigen (CEA) have been widely used in clinic, but the above disease markers are far from meeting the needs for diagnosing cancers at the early stage. The main reasons are as follows: (1) the sensitivity and specificity of the disease markers mentioned above are relatively low, therefore the detection results may yet not be taken as indicators for definite diagnosis of diseases; (2) the diagnosis rate of diseases at an early stage is in positive correlation with the therapeutic efficacy, but any of the disease markers mentioned above satisfies the requirement to diagnosis a disease at an early stage. Take cancer for instance, due to drawbacks including the extremely high specificity among types of differentiated cancers, the relatively low overall sensitivity of tumor bodies, the difficulty of repetitive sampling and the high requirements of sample preservation, and high costs, the application of current existing tumor markers cannot be widely promoted under the current conditions. Meanwhile, some conventional medical methods such as biopsy have inherent deficiencies, such as sampling at wrong sites, insufficiency of tissue and cell sample materials or lack of experience for the technician may all contribute to misdiagnosis. Other techniques like medical imaging that have been widely used in disease examination and diagnosis are rather limited in the qualitative description of disease process. Therefore, it is necessary to find novel disease markers that are sensitive and convenient in application, and capable of overcoming the defects of the currently existing markers.
Micro ribonucleic acids, also called miRNA, are a class of non-coding, single strand, small ribonucleic acid molecules with 19-23 nucleotides in length. They are highly conserved in evolution and closely related to many ordinary biophysical activities of animals, such as ontogenesis, tissue differentiation, apoptosis and energy metabolism, and are closely related to the occurrence and progression of many diseases. Recent research has shown that the expression levels of several miRNAs in chronic lymphocytic leukemia and Burkett lymphoma are down-regulated to varying extents (Lawrie C H, Gal S, Dunlop H M et al. Detection of elevated levels of tumor-associated miRNAs in serum of patients with diffuse large B-cell lymphoma. Br J Haematol 2008; 141:672-675); in the analysis of the expression of miRNAs in human lung cancer and breast cancer tissues, it was observed that the expression levels of some tissue-specific miRNAs is different from those of normal tissues (Garofalo M, Quintavalle C, Di Leva G et al. MicroRNA signatures of TRAIL resistance in human non-small cell lung cancer. Oncogene 2008). There are also researches proving that miRNAs affects the occurrence and progression of angiocardiopathies including cardiac hypertrophy, heart failure and atherosclerosis, and are closely related to metabolic diseases like diabetes type II (Tryndyak V P, Ross S A, Beland F A, Pogribny I P. Down-regulation of the microRNAs miR-34a, miR-127, and miR-200b in rat liver during hepatocarcinogenesis induced by a methyl-deficient diet. Mol Carcinog. 2008 Oct. 21). All these experimental results suggest that the expression and specific changes of miRNAs are definitely related to the occurrence and progression of diseases.
MiRNAs play a highly important role in the regulation of gene translation subsequent to the transcription, which indicates a relationship between miRNAs and diseases. Firstly, the change of miRNAs may be the cause of the diseases, because the inhibition and promotion factors of diseases may be targets of miRNAs. When the expression of miRNA itself is in disorder, for example when miRNAs that inhibit the disease promotion factors are down-regulated in expression, or miRNAs that inhibit the disease inhibition factors are up-regulated in expression, as a final result, the expression of a series of genes down-stream will be altered, and some gene-pathway will be in disorder, eventually causing diseases. Secondly, the changes of miRNAs are possibly the result of the diseases. This is because when a disease (cancers, for instance) occurs, it may cause loss of chromosome fragments, genetic mutation or instant amplification of chromosome fragments; if the miRNAs are from the affected fragments, the expression levels of these miRNAs will be significantly altered. Therefore, theoretically, miRNAs can be used as a novel class of disease markers; the specific alteration is undoubtedly related to disease occurrence and development. Meanwhile, miRNAs may be used as potential drug target sites; the occurrence and progression of the diseases may be largely relieved by inhibiting the up-regulated miRNAs or over-expressing the down-regulated miRNAs during the disease progression.
Presently, researches on miRNAs as disease markers have been carried out in China. For instance, Chinese patent applications CN100999765A and CN101298630A have disclosed respectively that, in the case of colon cancer, which ranks the 4th of malignant tumor occurrence rate, some miRNAs undergo specific alterations when benign colon polyps deteriorate into malignant tumor. A more sensitive and accurate method for diagnosing early-stage colon cancer has been developed according to the specific alterations of miRNAs. However, the wide clinical application is restrained by the difficulties in tissue sampling.